Abstract
Purpose: To purify β-galactosyltransferase, which is involved in the biosynthesis of keratan sulfate, from 2-day-old chick corneas. Methods: The activity was assayed using pyridylaminated GlcNAcβ1-3Galβ1-4Glc as acceptor substrate. Results: The β-galactosyltransferase did not bind to several ion exchange and affinity columns. In particular, it did not bind completely to an α-lactalbumin-agarose column. The partially purified enzyme showed an optimum pH at 7.0 [( N-[2-hydroxyethyl]piperazine- N′-[2-ethanesulfonic acid]) (HEPES) buffer]. Pyridylaminated GlcNAcβ1-3Galβ1-4GlcNAc, as well as the pyridylaminated GlcNAcβ1-3Galβ1-4Glc, served as the acceptor substrate of the enzyme, but not p-nitrophenyl-β-GlcNAc. The crude extract from chick corneas contained a high activity of the β-galactosyltransferase, which transfers Gal to pyridylaminated 6-sulfo-GlcNAcβ1-3Gal, but the activity was almost all lost during the purification procedures. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the enzyme fraction showed many bands. When each of the five main band proteins was partially analyzed for the amino acid sequence, none showed homology with the recently reported chick β-galactosyltransferases 1 and 2. Conclusions: β-Galactosyltransferase, which transfers Gal to 6-sulfo-GlcNAc end, was identified in chick corneas. The chick corneal β-galactosyltransferase(s) may be a novel one.
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